The formation of skeletal muscle during embryonic development and following injury to adult muscle requires fusion of myoblasts to form multinucleated myofibers. Recently, we discovered a novel muscle-specific membrane protein, named Myomaker, that controls vertebrate myoblast fusion during embryogenesis and adulthood. Myomaker is expressed on the cell surface of embryonic myoblasts during fusion and is down- regulated thereafter. Similarly, Myomaker is up-regulated in muscle satellite cells in response to injury, concomitant with their fusion during muscle regeneration. Over-expression of Myomaker in myoblasts dramatically enhances fusion and forced expression in fibroblasts promotes fusion with myoblasts. Conversely, genetic disruption of Myomaker in mice causes perinatal death due to an absence of multi- nucleated muscle fibers and conditional gene deletion in adult satellite cells completely prevents muscle regeneration. The discovery of Myomaker provides a new inroad into myoblast fusion and will enable the detailed molecular dissection of the mechanistic basis of this process. Myomaker belongs to a small family of related membrane proteins that are expressed in other cell types, suggesting a general mechanism for cell-cell fusion. The goals of this project are to define the precise molecular mechanism whereby Myomaker drives myoblast fusion and to identify additional components of the process through which Myomaker exerts its fusogenic activity. The insights gleaned from these studies will shed light not only on the fundamental mechanisms of intercellular fusion but will also have important implications for understanding muscle disease and for the potential development of new therapeutic strategies for restoration of function to diseased muscle.